New records of Trichinella patagoniensis from Argentina.

Trichinella spiralis was long considered the sole Trichinella species in Argentina. However, since 2004, various Trichinella species, including the encapsulated Trichinella patagoniensis and Trichinella britovi, as well as the unencapsulated Trichinella pseudospiralis, have been detected in the country. The present study aimed to identify Trichinella ML at the species level from cougars naturally infected from Argentina. To this end, muscle tissue samples from one cougar each from Córdoba, Neuquén, and Santa Cruz Provinces were individually analysed using the artificial digestion technique. DNA extraction and molecular identification of Trichinella species were conducted on individual muscle larvae by PCR amplification of the ESV region and subsequent PCR amplification and sequencing of the COI gene. Morphological analysis revealed muscle larvae with characteristics consistent with Trichinella genus. PCR revealed a single band of approximately 127 bp for each individual muscle larva. PCR amplification of the COI gene from each isolate generated a 309 bp band. Sequencing of the mitochondrial COI gene confirmed the identity of the parasite as T. patagoniensis. The present study documents new occurrences of T. patagoniensis in Puma concolor from Argentina, including the first detection of T. patagoniensis in Puma concolor from Córdoba and Neuquén Province. These findings expand the limited knowledge of T. patagoniensis distribution in Argentina.

Study of Trichinella patagoniensis in wild boars.

Trichinellosis is a zoonotic disease, which represents a significant public health concern in some South American countries, such as Argentina and Chile. Its impact is essentially due to absence of adequate control measures on meat from game animals, as well as the presence of illegal slaughterhouses and the trade of meat products without being tested for this parasite. In Argentina, trichinellosis is an endemic disease. At present, Trichinella spiralis, Trichinella patagoniensis, Trichinella pseudospiralis, and Trichinella britovi have been detected in animals from Argentina. Until now, T. patagoniensis had only been found in mountain cougars (Puma concolor) in Argentina but there is limited information available. The present study intends to determine susceptibility, serological response and distribution of muscle larvae in wild boars infected with T. patagoniensis, T. spiralis and T. pseudospiralis. For each of the Trichinella species five wild boars were inoculated with 20,000 muscle larvae. Except for two specimens which died during the experiment, the animals were euthanized 19 weeks post infection (pi). Blood samples were collected throughout the study in order to determine the antibody kinetics. Also, nine muscle samples from each specimen were taken and analysed for determination of larval distribution. Additionally, four muscle samples were used to obtain muscle juices. Wild boars infected with T. patagoniensis showed little to no larvae in the muscle samples analysed while animals infected with T. spiralis and T. pseudospiralis had a significantly high larval load in all the samples analysed. Optical density (OD) values remained above the cut-off value throughout the experiment. This is the first study to characterize the biological aspects of T. patagoniensis in wild boars.

Trichinella spiralis in a South American sea lion (Otaria flavescens) from Patagonia, Argentina.

Trichinella spp. from a sylvatic cycle has been found in several animal species such as pumas (Puma concolor), armadillos (Chaetophractus villosus), rats (Rattus norvegicus), and wild boars (Sus scrofa) in Argentina. Moreover, Trichinella infection has been detected in a wide range of marine mammals around the world, including polar bears (Ursus maritimus) and walruses (Odobenus rosmarus). Until the present time, Trichinella spp. infection has not been detected in marine mammals of South America. Samples from four South American sea lions (Otaria flavescens) found dead in Rio Negro, Argentina, were analyzed by artificial digestion, and in the case of one animal, Trichinella larvae were identified at the species level by nested multiplex PCR as Trichinella spiralis. This is the first report of a Trichinella species infecting marine mammals from South America.

Toxocariasis in Carnivora from Argentinean Patagonia: Species molecular identification, hosts, and geographical distribution.

Twenty four specimens of seven species belonging to the families Felidae, Mustelidae, and Canidae were obtained in Lanín and Nahuel Huapi National Parks from March 1996 to April 2016. Specimens were processed by necropsy in order to contribute to the knowledge of toxocariasis in wild carnivores of Argentinean Patagonia. The only Puma concolor and the seven Leopardus geoffroyi were positive for Toxocara cati. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) of the ITS-1 region from larval and adult DNA was carried out to confirm parasite species identification. This is the first molecular determination of T. cati from wild felids in Argentina and the study also fill gaps about the spatial distribution and hosts for Toxocara cati.

Evaluation of the infectivity and the persistence of Trichinella patagoniensis in muscle tissue of decomposing guinea pig (Cavia porcellus).

Trichinella patagoniensis, a new species of Trichinella, is widespread in Argentina. The success of parasite transmission depends, among other factors, on the resistance of L1 larvae present in the muscle tissue (ML) of dead hosts undergoing the decomposition process in different environmental conditions. The aim of the present work was to study the infectivity of T. patagoniensis muscle larvae in Cavia porcellus and the capability of the parasite to survive in decomposed muscle tissue of guinea pigs subjected to different environmental conditions. Thirty-two female Ssi:AL guinea pigs were orally inoculated with 2000 ML of T. patagoniensis (ISS2311). All the animals were sacrificed 42 days post-infection. Twenty-six animals were eviscerated, and carcasses were placed on the surface of soil inside plastic boxes that were exposed to environmental conditions in the summer 2014-2015 and autumn of 2015 in Buenos Aires, Argentina. Carcasses from six animals were placed into a plastic box inside the refrigerator at a temperature of 4 °C. The muscle tissue samples from the carcasses were examined weekly for the presence of larvae, and the infectivity of recovered ML was tested in BALB/c mice. Our results showed for the first time the ability of T. patagoniensis to complete its life cycle in guinea pigs, thus serving as a potential natural host. Also, larvae of T. patagoniensis remained infective in muscle tissue for several weeks while undergoing decomposition under different environmental conditions.

Susceptibility of chickens (Gallus gallus domesticus) to Trichinella patagoniensis.

Trichinella spp. is a genus of parasites that is widespread all over the world. In Argentina, T. spiralis was for years the only species involved in human and animal outbreaks. During the last decade, T. patagoniensis, a new Trichinella species, was discovered in Argentina. Up to now, this species has only been found in cougars (Puma concolor). Experimental infections in pigs, cats, mice and rats with this new genotype showed that cats and mice were the most susceptible hosts. The aim of the present work was to evaluate the susceptibility of chickens to infection with T. patagoniensis. In order to study the intestinal and muscular phase, and the histopathological changes, 27 Leghorn chickens were inoculated per os with 1000 muscle larvae of T. patagoniensis and were euthanized on days 4, 5, 6, 7, 11, 14, 21, 28 and 35. Adult worms of T. patagoniensis were recovered from the small intestine of chickens up to day 14p.i. Gross examination of small intestine showed a moderate congestive appearance. Microscopically, an inflammatory response with lymphocytes and eosinophils in lamina propria, slight hyperemia, oedema and some haemorrhagic areas were detected. Lesions observed in chickens were similar to those described in different animal species during the intestinal phase. No muscular larvae were recovered from the muscle samples. These results suggest that T. patagoniensis is not capable to complete its entire life cycle in chickens.

Experimental infection with Trichinella T12 in domestic cats.

Trichinella spiralis has been documented in wild animals in Argentina, including puma, armadillos, rats and wild boars. In 2008, molecular analysis identified Trichinella T12 from a naturally infected puma (Puma concolor) from Patagonia. The aim of the present work was to study the relationship between the infectivity and pathology of Trichinella T12 in the puma and in domestic cats, and the possible risks that may be present for transmission among these animals. Two cats (A and B) were orally-infected with 3300 and 1850 Trichinella T12 muscle larvae, respectively; one additional cat was used as a control. During the 54 days post-infection, a daily examination was performed which included monitoring body temperature, and cardiac and respiration rates; the animals were then euthanized. Hematological studies included hematocrit (%), hemoglobin (g/dl), and white cell, neutrophil, lymphocyte and eosinophil counts. Blood biochemistry included urea, creatinine, AST, ALT, CK, LDH and ALP. An ELISA assay was also performed. At necropsy, organs (liver, spleen, brain, cerebellum and kidney), nails and muscle samples were obtained for histopathology studies and artificial digestion. The muscles that were studied included the diaphragm, massetter, cutaneous, temporal, intercostals, lumbar, tongue, limbs, neck and tail. Clinical signs, such as anorexia, diarrhea, vomiting, shaggy hair, decay and muscle pain, were observed in both cats. The eosinophil counts were elevated in both cats A and B. Trichinella larvae were recovered from all of the muscles analyzed where the histopathology showed larvae in several muscles without degenerative reaction. Neither larvae nor lesions were observed in non-muscular organs. Cat A had a maximum of 246 larvae per gram (lpg) in the temporal muscle and a minimum of 80 lpg in the tongue, while cat B had a maximum of 65 lpg in muscles of the leg and a minimum of 10 lpg in tail muscles. This study represents the first record of experimental infection of cats with Trichinella T12.

Epidemiological survey of Trichinella infection in domestic, synanthropic and sylvatic animals from Argentina.

The presence of Trichinella larvae was investigated in 247 samples taken from domestic, synanthropic and sylvatic animals, collected during 1996 to 2005 in 12 endemic provinces of Trichinella infection in Argentina. Muscle larvae of Trichinella from 65 infected animals were identified at the species level by single larva nested polymerase chain reaction (PCR) technique based on the variability within the expansion segment V (ESV) region of the ribosomal DNA. Trichinella infections were found in 97 of 164 pigs, 38 of 56 pork products, two domestic dogs, one domestic cat, 7 of 11 armadillos and 3 of 9 synanthropic rats. All Trichinella isolates were identified as Trichinella spiralis by nested PCR. These findings add new data on the epidemiology of trichinellosis and should be considered when implementing new strategies to control this zoonosis.

Porcine and rodent infection with Trichinella, in the Sierra Grande area of Rio Negro province, Argentina.

In 2000, two cases of human trichinellosis were detected in the Sierra Grande area of Rio Negro province, Argentina. As part of an investigation of the aetiology of these cases, 300 pigs slaughtered for consumption in the area between 2000 and 2002 were checked for Trichinella infection, by artificial digestion of a muscle sample. Twelve (5.6%) - four (7.3%) of the 55 checked in 2000, five (4.8%) of the 105 investigated in 2001, and three (2.1%) of the 140 investigated in 2002 - were found infected. Blood samples were collected from other pigs aged > 6 months old, so that sera could be tested, in ELISA and by western blotting, for anti- Trichinella antibodies. Of the 181 animals checked in the initial serological survey, 36 (19.9%) were found seropositive for Trichinella. When 35 of the seronegative pigs were re-checked 6 months later, three (8.6%) were found to have seroconverted. Four (15.4%) of 26 local rodents, caught in Sherman-type traps, were also found positive when checked for infection by artificial digestion. It appears that about 20% of pigs in the study area are infected each year, this high level of transmission being sustained by a high prevalence of infection in the local rodent populations.